Drug addiction has. been characterizedas a progressivealteration of the brain reward system. Using animals to model this progression, it is clear that each stage of addiction,from initial response to relapse, is strongly influenced by genetic contributions [unreadable]The collection of genes that can potentiallyregulate this process is quite large, and it has become a central question of drug abuse research to identify the key molecular determinants for genetic susceptibility and therapeutic interventidn. These genes can be identified by classical genetics, which has been a widely successful method to uncover key components of biological pathwaysand regulators of complex behaviors. As an unbiased approach it affords the opportunity to discover novel genes that might not be otherwise implicated in addiction. This proposal describes a genetic screen to identify genes that regulate dopamine neurotransmission. Because dopaminergic pathways are strongly implicated in reward mechanisms, these genes may also play a role in the process of addiction. The proposed approach is a genome-wide saturation mutagenesis effort and phenotypic screen that is tailored to select for dopamine-modulated behaviors. Saturation mutagenesis is achieved in the mouse with the alkylating agent ethylnitrosourea (ENU), which induces base pair mutations at high frequency. These mutations are then bred onto a genetic background of dopfamine transporter (DAT) deficiency. DAT knockout mice lack the ability to clear synaptic dopamine, which results in persistent hyperdopaminergia. The phenotypic screen is designed to find new mutations that enhance or suppress the behavioral phenotype of DAT knockout mice. It is likely that these modifier mutations will directly or indirectly influence dopamine transmission.The research plan has the following aims: AIM I. To introduce random mutations onto DAT +/- and -/- genetic backgrounds. AIM II. To establish and implement a forward screen to find dominant modifiers of the DAT mutant phenotype. AIM III. To determine the identity of novel DAT modifier mutations through chromosome positioning and candidate gene analysis. AIM IV. To characterize the neurochemistry and behavioral pharmacology of mutant mouse lines, and determine the influence of the mutation on dopamine neurotransmission. The ultimate goal of the research plan is to isolate new alleles that enhance or impair dopamine signals, and determine whether these alleles play a role in addiction or could be targets for therapeutic treatments of human addictions. This research proposal has also been designed to further develop the scientific career of the P.I. in preparation for an independent faculty position using forward genetic approaches to dissect neurotransmitter signaling pathways and complex behaviors related to human disease states.